A cell wall lytic enzyme is an enzyme degrading the cell wall of bacteria including Actinomycetes, and on the action of the enzyme, the bacterial cell wall is decomposed, leading to the death of the bacteria.
The outer layer of bacteria is covered with cell wall, and the principal structural component of the cell wall is a peptide glycan comprising sugar chains and peptides. The cell wall lytic enzyme acts on the peptide glycan.
When the enzyme acts on the peptide glycan, the enzyme reacts with the sugar chain of the peptide glycan to generate N-acetylmuramic acid from the sugar at the terminus to be reduced. Therefore, the enzyme is classified as N-acetylmuramidase.
Major enzymes to be classified as N-acetylmuramidase include lysozyme derived from chicken egg white. However, the cell wall lytic enzyme is different from these enzymes in terms of enzymatic and chemical properties and the subjective microorganisms to be decomposed [Hayashi K., et al., Agric. Biol. Chem. (European Edition of Japanese Journal of Agriculture, Biochemistry and Chemistry), Vol. 45, pp. 2289-2300, 1981], and the enzyme has novel specificities.
As has been described above, the cell wall lytic enzyme decomposes bacterial cell wall, and by utilizing the property, the enzyme is used for extracting enzymes and DNA present in the inside of bacteria.
Because some bacteria may be killed through the action of the present enzyme, furthermore, the enzyme may be utilized as food preservative [Hayashi K., et al., Agric. Biol. Chem. (European Edition of Japanese Journal of Agriculture, Biochemistry and Chemistry), Vol. 53, pp. 3173-3177, 1989].
Conventional methods for recovering the cell wall lytic enzyme include a method comprising culturing microorganisms, such as Actinomycetes belonging to genus Streptomyces, and bacteria belonging to genera Achromobacter, Aeromonas, Bacillus, Clostridium, Flavobacterium, Myxobacter, Myxococcus, Pseudomonas, Staphylococcus and Streptococcus, and preparing the objective enzyme from the culture filtrate or the cultured bacteria. When the cell wall lytic enzyme is egg white-derived lysozyme, use is made of a method comprising preparing the enzyme by utilizing isoelectric precipitation and the like.
The enzyme recovered by these methods is commercially available as crude enzyme or purified enzyme. However, these methods are not satisfactory as methods for producing the enzyme in a stable fashion.
For further promotion of the utilization of cell wall lytic enzymes, an object of the present invention resides in making contribution to the industrial production of cell wall lytic enzymes, by cloning the gene of said enzymes to elucidate the structure of the gene and expressing said gene.